Dosing regimens for 2-hydroxy-6-((2-(1-isopropyl-1h-pyrazol-5-yl)pyridin-3-yl)methoxy)benzaldehyde

ABSTRACT

Provided herein are methods for treating sickle cell disease, comprising administering to a subject 2-hydroxy-6-((2-(1-isopropyl-1H-pyrazol-5-yl)pyridin-3-yl)-methoxy)benzaldehyde (Compound 1), or a polymorph thereof, in certain dosing regimens.

1. CROSS-REFERENCE TO RELATED APPLICATIONS

The present application claims the benefit of priority of U.S.Provisional Application No. 62/263,554, filed Dec. 4, 2015, and U.S.Provisional Application No. 62/375,832, filed Aug. 16, 2016, the contentof each which is hereby incorporated by reference in its entirety.

2. FIELD OF THE INVENTION

Provided herein are compounds, compositions, formulations, dosage formsand methods for the treatment of sickle cell disease. As providedherein, such treatment may comprise administering to a subject, orpreparing for administration to such subject,2-hydroxy-6-((2-(1-isopropyl-1H-pyrazol-5-yl)pyridin-3-yl)-methoxy)benzaldehyde,or a polymorph thereof, in certain dosing regimens. Also provided hereinis a capsule dosage form comprising high drug loads of2-hydroxy-6-((2-(1-isopropyl-1H-pyrazol-5-yl)pyridin-3-yl)methoxy)benzaldehydeor a polymorph thereof.

3. BACKGROUND OF THE INVENTION

Hemoglobin (Hb) is a tetrameric protein in red blood cells thattransports up to four oxygen molecules from the lungs to various tissuesand organs throughout the body.

Hemoglobin binds and releases oxygen through conformational changes, andis in the tense (T) state when it is unbound to oxygen and in therelaxed (R) state when it is bound to oxygen. The equilibrium betweenthe two conformational states is under allosteric regulation. Naturalcompounds such as 2,3-bisphosphoglycerate (2,3-BPG), protons, and carbondioxide stabilize hemoglobin in its de-oxygenated T state, while oxygenstabilizes hemoglobin in its oxygenated R state. Other relaxed R stateshave also been found, however their role in allosteric regulation hasnot been fully elucidated.

Sickle cell disease is a prevalent disease particularly among those ofAfrican and Mediterranean descent. Sickle hemoglobin (HbS) contains apoint mutation where glutamic acid is replaced with valine, allowing theT state to become susceptible to polymerization to give the HbScontaining red blood cells their characteristic sickle shape. Thesickled cells are also more rigid than normal red blood cells, and theirlack of flexibility can lead to blockage of blood vessels. Certainsynthetic aldehydes have been found to shift the equilibrium from thepolymer forming T state to the non-polymer forming R state (Nnamani etal., Chemistry & Biodiversity Vol. 5, 2008 pp. 1762-1769) by acting asallosteric modulators to stabilize the R state through formation of aSchiff base with an amino group on hemoglobin.

U.S. Pat. No. 7,160,910 discloses 2-furfuraldehydes and relatedcompounds that are also allosteric modulators of hemoglobin. Oneparticular compound, 5-hydroxymethyl-2-furfuraldehyde (5HMF), was foundto be a potent hemoglobin modulator both in vitro and in vivo. 5HMF iscurrently in clinical trials for treatment of sickle cell disease.However, 5HMF requires 4 times daily dosing of 1,000 mg (see, e.g.,ClinicalTrials.gov; NCT01987908). This requirement for frequent dosingat relatively high amounts can present problems with patient complianceand high treatment costs.

Accordingly, there exists a need for effective methods of treatingsickle cell disease, which use compounds that are effective whenadministered at lower doses.

4. SUMMARY

Applicant has unexpectedly found that Compound 1 disclosed herein istherapeutically effective in the treatment of sickle cell disease (SCD)at low doses, in spite of the large concentration of hemoglobin in redcells (5 nM in red cells).

In one aspect, provided herein are methods for treating sickle celldisease in a patient comprising administering to the patient Compound 1:

wherein the compound is administered in a dose of from about 500 mg/dayto about 1500 mg/day. In one embodiment of the first aspect, Compound 1is administered in a dose of about 1100, about 1200, about 1300, about1400, or about 1500 mg/day. In another embodiment of the first aspect,Compound 1 is administered in a dose of about 1050, about 1100, about1150, about 1200, about 1250, about 1300, about 1350, about 1400, about1450, or about 1500 mg/day. In another embodiment of the first aspect,the compound is administered in a dose of about 500, about 550, about600, about 650, about 700, about 750, about 800, about 850, about 900,about 950, about 1000, about 1050, about 1100, about 1150, about 1200,about 1250, about 1300, about 1350, about 1400, about 1450, or about1500 mg/day. In another embodiment of the first aspect, the compound isadministered in a dose of from about 500 mg/day to about 1000 mg/day. Inanother embodiment of the first aspect, the compound is administered ina dose of about 500, about 550, about 600, about 650, about 700, about750, about 800, about 850, about 900, about 950, or about 1000 mg/day.In another embodiment of the first aspect, the compound is administeredin a dose of about 600, about 650, about 700, about 750, about 800,about 850, or about 900 mg/day. In yet another embodiment of the firstaspect, the compound is administered in a dose of from about 500 mg/dayto about 900 mg/day. In yet another embodiment of the first aspect, thecompound is administered in a dose of from about 600 mg/day to about 900mg/day. In yet another embodiment of the first aspect the compound isadministered in a dose of about 700 mg/day. In yet another embodiment ofthe first aspect, the compound is administered in a dose of about 600mg/day. In yet another embodiment of the first aspect, the compound isadministered in a dose of about 900 mg/day. In yet another embodiment ofthe first aspect, the compound is administered in a dose of about 1200mg/day. In yet another embodiment of the first aspect, the compound isadministered in a dose of about 1500 mg/day. In yet another embodimentof the first aspect, the compound is administered in a dose of 900mg/day. In yet another embodiment of the first aspect, the compound isadministered in a dose of 1200 mg/day. In yet another embodiment of thefirst aspect, the compound is administered in a dose of 1500 mg/day. Inyet another embodiment of the first aspect and embodiments containedtherein, the patient is in need to treatment.

In a second embodiment of the first aspect and embodiments containedtherein above, the compound is administered once daily.

In a third embodiment of the first aspect and embodiments containedtherein above (which include the second embodiment), the dose isadministered in a capsule or tablet. Within the third embodiment, in onesubembodiment, the dose is administered in a 100 mg or a 300 mg capsule.Within the third embodiment, in another subembodiment, the dose isadministered in a 300 mg capsule.

In a fourth embodiment of the first aspect and embodiments containedtherein above (including the second and third embodiments andsubembodiments contained therein), Compound 1 is a crystalline ansolvateform. In one embodiment, the crystalline ansolvate is Form IIcharacterized by at least one X-ray powder diffraction peak (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In one embodiment, the crystalline ansolvate is characterizedby at least one X-ray powder diffraction peak (Cu Kα radiation) selectedfrom 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In anotherembodiment, Form II is characterized by at least two X-ray powderdiffraction peaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ). In another embodiment, the crystallineansolvate is characterized by at least two X-ray powder diffractionpeaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ). In yet another embodiment, Form II ischaracterized by at least three X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In yet another embodiment, the crystalline ansolvate ischaracterized by at least three X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In yet another embodiment, Form IT is characterized by X-raypowder diffraction peaks (Cu Kα radiation) of 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ). In yet another embodiment, the crystallineansolvate is characterized by X-ray powder diffraction peaks (Cu Kαradiation) of 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yetanother embodiment, Form II is characterized by an X-ray powderdiffraction pattern (Cu Kα radiation) substantially similar to that ofFIG. 1. In yet another embodiment, the crystalline ansolvate ischaracterized by an X-ray powder diffraction pattern (Cu Kα radiation)substantially similar to that of FIG. 1. In yet another embodiment, thecrystalline ansolvate form of Compound 1 is substantially free of Form Iand/or Form N. Form I of Compound 1 is characterized by at least threeX-ray powder diffraction peaks (Cu Kα radiation) at 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andForm N of Compound 1 is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) at 11.65°, 11.85°, 12.08°, 16.70°,19.65° and 23.48°2θ (each ±0.2°2θ).

In a second aspect, provided is a method of treating interstitialpulmonary fibrosis in a patient comprising administering to the patientabout 1100 mg/day to about 1500 mg/day of Compound 1 optionally incombination with an anti-fibrotic agent. In one embodiment, theanti-fibrotic agent is selected from pirfenidone, nintenabib, andsystemic corticosteroids.

In one embodiment of the second aspect, Compound 1 is administered in adose of about 1100, about 1200, about 1300, about 1400, or about 1500mg/day. In another embodiment of the second aspect, Compound 1 isadministered in a dose of about 1050, about 1100, about 1150, about1200, about 1250, about 1300, about 1350, about 1400, about 1450, orabout 1500 mg/day. In another embodiment of the second aspect, thecompound is administered in a dose of about 1200 mg/day. In anotherembodiment of the second aspect, the compound is administered in a doseof about 1500 mg/day. In another embodiment of the second aspect, thecompound is administered in a dose of 1200 mg/day. In another embodimentof the second aspect, the compound is administered in a dose of 1500mg/day. In yet another embodiment of the second aspect and embodimentscontained therein, the patient is in need to treatment.

In a second embodiment of the second aspect and embodiments containedtherein above, the compound is administered once daily.

In a third embodiment of the second aspect and embodiments containedtherein above (which include the second embodiment), the compound isadministered in a capsule or tablet. Within the third embodiment, in onesubembodiment, the compound is administered in a 100 mg or a 300 mgcapsule. Within the third embodiment, in another subembodiment, thecompound is administered in a 300 mg capsule.

In a fourth embodiment of the second aspect and embodiments containedtherein above (including the second and third embodiments andsubembodiments contained therein), Compound 1 is a crystalline ansolvateform. In one embodiment, the crystalline ansolvate is Form IIcharacterized by at least one X-ray powder diffraction peak (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In one embodiment, the crystalline ansolvate is characterizedby at least one X-ray powder diffraction peak (Cu Kα radiation) selectedfrom 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In anotherembodiment, Form II is characterized by at least two X-ray powderdiffraction peaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ). In another embodiment, the crystallineansolvate is characterized by at least two X-ray powder diffractionpeaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ). In yet another embodiment, Form II ischaracterized by at least three X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (eachf0.2°2θ). In yet another embodiment, the crystalline ansolvate ischaracterized by at least three X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In yet another embodiment, Form II is characterized by X-raypowder diffraction peaks (Cu Kα radiation) of 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ). In yet another embodiment, the crystallineansolvate is characterized by X-ray powder diffraction peaks (Cu Kαradiation) of 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yetanother, Form II is characterized by an X-ray powder diffraction pattern(Cu Kα radiation) substantially similar to that of FIG. 1. In yetanother, the crystalline ansolvate is characterized by an X-ray powderdiffraction pattern (Cu Kα radiation) substantially similar to that ofFIG. 1.

In a third aspect, provided is a capsule dosage form comprising:

(i) from about 65% to about 93% w/w of Compound 1 or a polymorphthereof; and

(ii) from about 2% to about 10% w/w a binder;

wherein w/w is relative to the total weight of the formulation(excluding the weight of the capsule). With regards to the capsuleformulation; “about” means ±1°% of a given range or value.

In one embodiment of the third aspect, the capsule dosage form furthercomprises from about 2% to about 10% a disintegrant.

In a second embodiment of the third aspect, the capsule dosage formfurther comprises from about 2% to about 10% a disintegrant and about 2%to 35% a filler.

In a fourth aspect, provided is a capsule dosage form comprising:

(i) from about 65% to about 86% w/w of Compound 1 or a polymorphthereof;

(ii) from about 2% to about 6% w/w a binder;

(iii) from about 6% to about 25% w/w a filler;

(iv) from about 2% to 6% w/w a disintegrant; and

(iv) from about 0.5% to about 1.5% w/w a lubricant;

wherein w/w is relative to the total weight of the formulation(excluding the weight of the capsule). With regards to the capsuleformulation; “about” means ±1°% of a given range or value.

In one embodiment of the fourth aspect, the capsule dosage formcomprises:

(i) from about 65% to about 86% w/w of Compound 1 or a polymorphthereof;

(ii) from about 2% to about 6% w/w a binder;

(iii) from about 3.5% to about 25% w/w an insoluble filler or 2.5% to25% w/w of soluble filler or 2.5% to 25% of a combination of soluble orinsoluble filler;

(iv) from about 2% to 6% w/w a disintegrant; and

(iv) from about 0.5% to about 1.5% w/w a lubricant.

In a second embodiment of the fourth aspect, the capsule dosage formcomprises:

(i) about 86% w/w of Compound 1 or a polymorph thereof;

(ii) about 4% w/w a binder;

(iii) about 3.5% w/w an insoluble filler and 2.5% w/w of soluble filler;

(iv) about 3.5% w/w a disintegrant; and

(iv) about 0.5% w/w a lubricant.

In a third embodiment of the fourth aspect, the capsule dosage formcomprises:

(i) 85.71% w/w of Compound 1 or a polymorph thereof;

(ii) 4% w/w a binder;

(iii) 3.64% w/w an insoluble filler and 2.65% w/w of soluble filler;

(iv) 2.65% w/w a disintegrant; and

(iv) 0.5% w/w a lubricant.

In one embodiment of the third and fourth aspects, and embodimentscontained therein:

Compound 1 is Form II substantially free of Form I and/or N;

the binder is hypromellose;

the insoluble filler is microcrystalline cellulose

the soluble filler is lactose monohydrate;

the disintregrant is croscarmellose sodium; and

the lubricant is magnesium stearate.

In another embodiment of the third and fourth aspects, and embodimentscontained therein, the capsule contains 300 mg of Compound 1 Form IIsubstantially free of Form I and/or N.

In another embodiment of the third and fourth aspects, and embodimentscontained therein, Compound 1 is a crystalline ansolvate form. In oneembodiment, the crystalline ansolvate is Form II characterized by atleast one X-ray powder diffraction peak (Cu Kα radiation) selected from13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In one embodiment,the crystalline ansolvate is characterized by at least one X-ray powderdiffraction peak (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ). In another embodiment, Form II ischaracterized by at least two X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In another embodiment, the crystalline ansolvate ischaracterized by at least two X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In yet another embodiment, Form II is characterized by atleast three X-ray powder diffraction peaks (Cu Kα radiation) selectedfrom 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yet anotherembodiment, the crystalline ansolvate is characterized by at least threeX-ray powder diffraction peaks (Cu Kα radiation) selected from 13.37°,14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yet another embodiment,Form IT is characterized by X-ray powder diffraction peaks (Cu Kαradiation) of 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yetanother embodiment, the crystalline ansolvate is characterized by X-raypowder diffraction peaks (Cu Kα radiation) of 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ). In yet another, Form II is characterized by anX-ray powder diffraction pattern (Cu Kα radiation) substantially similarto that of FIG. 1. In yet another, the crystalline ansolvate ischaracterized by an X-ray powder diffraction pattern (Cu Kα radiation)substantially similar to that of FIG. 1.

In another embodiment of the third and fourth aspects, and embodimentscontained therein, the capsule contains 300 mg±5% of Compound 1, whereincompound 1 is a crystalline ansolvate form that is characterized by atleast two X-ray powder diffraction peaks (Cu Kα radiation) selected from13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ); wherein thecrystalline ansolvate form is substantially free of Form I and/or N;wherein Form I is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andwherein Form N is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 11.65°, 11.85°,12.08°, 16.70°, 19.65° and 23.48°2θ (each ±0.2°2θ).

Due to the high drug loading, higher doses of Compound 1 can bedelivered with minimal number of dosing units making it practical from aconvenience, compliance and marketing perspective. Additionally, inspite of high drug loading, the capsule formulation displays superiorphysical properties due to the appropriate ratio of the binder to thewet granulation process parameters. Further, the combination of solubleand insoluble fillers gives granule strength, flow properties anddisintegration that provides the desired therapeutic effect.

5. BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a XRPD profile and contemplated indexing for the free baseForm II anhydrous crystal of Compound 1.

FIG. 2 illustrates whole blood concentration at steady state for twodoses (500 mg, 700 mg) of Compound 1.

FIG. 3 illustrates representative oxygen equilibrium curves for twodoses (500 mg, 700 mg) of Compound 1, with comparison to placebo.

FIG. 4 illustrates change in hemoglobin (g/dL) over time for two doses(500 mg, 700 mg) of Compound 1, with comparison to placebo.

FIG. 5 illustrates percent (%) change in reticulocytes over time for twodoses (500 mg, 700 mg) of Compound 1, with comparison to placebo.

FIG. 6 illustrates percent (%) sickle cells over time for two doses (500mg, 700 mg) of Compound 1, with comparison to placebo.

FIGS. 7A-7B provide representative images of sickle cells from subjecttreated with 700 mg of Compound 1, over a period of one day as shown inFIG. 7A; and twenty-eight (28) days as shown in FIG. 7B.

FIG. 8 illustrates the percent (%) change in reticulocytes to day 28versus whole blood concentration of Compound 1.

FIGS. 9A-9D illustrate the linear relationship between Compound 1 wholeblood concentrations and effect on hemolytic measures: FIG. 9A showspercent (%) change in absolute reticulocytes; FIG. 9B shows percent (%)change in unconjugated bilirubin; FIG. 9C shows percent (%) change inLDH; and FIG. 9D shows percent (%) change in hemoglobin.

6. DETAILED DESCRIPTION OF THE INVENTION

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of ordinary skillin the art. All patents, applications, published applications and otherpublications are incorporated by reference in their entirety. In theevent that there is a plurality of definitions for a term herein, thosein this section prevail unless stated otherwise.

6.1 Definitions

As used herein, the below terms have the following meanings unlessspecified otherwise.

It is noted here that as used in this specification and the appendedclaims, the singular forms “a,” “an,” and “the” and the like includeplural referents unless the context clearly dictates otherwise.

The term “about” or “approximately” means an acceptable error for aparticular value as determined by one of ordinary skill in the art,which depends in part on how the value is measured or determined. Withregards to the dose, the term “about” or “approximately” means within 1,2, 3, or 4 standard deviations. In certain embodiments, the term “about”or “approximately” means within 50%, 30%, 20%, 15%, 10%, 9%, 8%, 7%, 6%,5%, 4%, 3%, 2%, 1%, 0.5%, or 0.05% of a given dose. In certainembodiments, the term “about” or “approximately” means within 50%, 20%,15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.05% of a dose.In certain embodiments, the term “about” or “approximately” means within0.5% to 1% of a given dose.

The term “administration” refers to introducing an agent into a patient.A therapeutic amount can be administered, which can be determined by thetreating physician or the like. An oral route of administration ispreferred. The related terms and phrases administering” and“administration of”, when used in connection with a compound orpharmaceutical composition (and grammatical equivalents), refer both todirect administration, which may be administration to a patient by amedical professional or by self-administration by the patient, and/or toindirect administration, which may be the act of prescribing a drug. Forexample, a physician who instructs a patient to self-administer a drugand/or provides a patient with a prescription for a drug isadministering the drug to the patient. In any event, administrationentails delivery to the patient of the drug.

The “crystalline ansolvate” of Compound 1 is a crystalline solid form ofthe free base of2-hydroxy-6-((2-(1-isopropyl-1H-pyrazol-5-yl)pyridin-3-yl)methoxy)benzaldehyde,such as, e.g., crystalline Form I, Form II or Material N as disclosed inInternational Publication No. WO 2015/120133 A1 (see, e.g., pages 3-9and pages 51-54), the disclosure of which is incorporated herein byreference in its entirety.

“Characterization” refers to obtaining data which may be used toidentify a solid form of a compound, for example, to identify whetherthe solid form is amorphous or crystalline and whether it is unsolvatedor solvated. The process by which solid forms are characterized involvesanalyzing data collected on the polymorphic forms so as to allow one ofordinary skill in the art to distinguish one solid form from other solidforms containing the same material. Chemical identity of solid forms canoften be determined with solution-state techniques such as ³C NMR or ¹HNMR. While these may help identify a material, and a solvent moleculefor a solvate, such solution-state techniques themselves may not provideinformation about the solid state. There are, however, solid-stateanalytical techniques that can be used to provide information aboutsolid-state structure and differentiate among polymorphic solid forms,such as single crystal X-ray diffraction, X-ray powder diffraction(XRPD), solid state nuclear magnetic resonance (SS-NMR), and infraredand Raman spectroscopy, and thermal techniques such as differentialscanning calorimetry (DSC), solid state ³C-NMR, thermogravimetry (TG),melting point, and hot stage microscopy.

To “characterize” a solid form of a compound, one may, for example,collect XRPD data on solid forms of the compound and compare the XRPDpeaks of the forms. For example, the collection of peaks whichdistinguish e.g., Form II from the other known forms is a collection ofpeaks which may be used to characterize Form IT. Those of ordinary skillin the art will recognize that there are often multiple ways, includingmultiple ways using the same analytical technique, to characterize solidforms. Additional peaks could also be used, but are not necessary, tocharacterize the form up to and including an entire diffraction pattern.Although all the peaks within an entire XRPD pattern may be used tocharacterize such a form, a subset of that data may, and typically is,used to characterize the form.

An XRPD pattern is an x-y graph with diffraction angle (typically °2θ)on the x-axis and intensity on the y-axis. The peaks within this patternmay be used to characterize a crystalline solid form. As with any datameasurement, there is variability in XRPD data. The data are oftenrepresented solely by the diffraction angle of the peaks rather thanincluding the intensity of the peaks because peak intensity can beparticularly sensitive to sample preparation (for example, particlesize, moisture content, solvent content, and preferred orientationeffects influence the sensitivity), so samples of the same materialprepared under different conditions may yield slightly differentpatterns; this variability is usually greater than the variability indiffraction angles. Diffraction angle variability may also be sensitiveto sample preparation. Other sources of variability come from instrumentparameters and processing of the raw X-ray data: different X-rayinstruments operate using different parameters and these may lead toslightly different XRPD patterns from the same solid form, and similarlydifferent software packages process X-ray data differently and this alsoleads to variability. These and other sources of variability are knownto those of ordinary skill in the pharmaceutical arts. Due to suchsources of variability, it is usual to assign a variability of ±0.2°2θto diffraction angles in XRPD patterns.

“Comprising” or “comprises” is intended to mean that the compositionsand methods include the recited elements, but not exclude others.“Consisting essentially of” when used to define compositions andmethods, shall mean excluding other elements of any essentialsignificance to the combination for the stated purpose. Thus, acomposition consisting essentially of the elements as defined hereinwould not exclude other materials or steps that do not materially affectthe basic and novel characteristic(s) of the claimed invention.“Consisting of” shall mean excluding more than trace elements of otheringredients and substantial method steps. Embodiments defined by each ofthese transition terms are within the scope of this invention.

The term “dose” or “dosage” refers to the total amount of activematerial (e.g., Compound 1 disclosed herein) administered to a patientin a single day (24-hour period). The desired dose may be administeredonce daily, for example, as a single bolus. Alternatively, the desireddose may be administered in one, two, three, four or more subdoses atappropriate intervals throughout the day, where the cumulative amount ofthe subdoses equals the amount of the desired dose administered in asingle day. The terms “dose” and “dosage” are used interchangeablyherein.

The term “dosage form” refers to physically discrete units, each unitcontaining a predetermined amount of active material (e.g., Compound 1disclosed herein) in association with the required excipients. Suitabledosage forms include, for example, tablets, capsules, pills, and thelike.

The capsule of the present disclosure comprises excipients such as apharmaceutically acceptable binder, filler (also known as diluent),disintegrant, and lubricant. Excipients can have two or more functionsin a pharmaceutical composition. Characterization herein of a particularexcipient as having a certain function, e.g., filler, disintegrant,etc., should not be read as limiting to that function. Furtherinformation on excipients can be found in standard reference works suchas Handbook of Pharmaceutical Excipients, 3rd ed. (Kibbe, ed. (2000),Washington: American Pharmaceutical Association).

A “disintegrant” as used herein refers to an excipient that can breakupor disintegrate the formulation when it comes in contact with, forexample, the gastrointestinal fluid. Suitable disintegrants include,either individually or in combination, starches including pregelatinizedstarch and sodium starch glycolate; clays; magnesium aluminum silicate;cellulose-based disintegrants such as powdered cellulose,microcrystalline cellulose, methylcellulose, low-substitutedhydroxypropylcellulose, carmellose, carmellose calcium, carmellosesodium and croscarmellose sodium; alginates; povidone; crospovidone;polacrilin potassium; gums such as agar, guar, locust bean, karaya,pectin and tragacanth gums; colloidal silicon dioxide; and the like. Inone embodiment, the disintegrant is carmellose sodium. In oneembodiment, the disintegrant is powdered cellulose, microcrystallinecellulose, methylcellulose, or low-substituted hydroxypropylcellulose,or a combination thereof. In one embodiment, the disintegrant iscarmellose, carmellose calcium, carmellose sodium or croscarmellosesodium, or a combination thereof. In one embodiment, the disintegrant iscroscarmellose sodium.

Lubricants as used herein refers to an excipient that reduces frictionbetween the mixture and equipment during granulation process. Exemplarylubricants include, either individually or in combination, glycerylbehenate; stearic acid and salts thereof, including magnesium, calciumand sodium stearates; hydrogenated vegetable oils; glycerylpalmitostearate; talc; waxes; sodium benzoate; sodium acetate; sodiumfumarate; sodium stearyl fumarate; PEGs (e.g., PEG 4000 and PEG 6000);poloxamers; polyvinyl alcohol; sodium oleate; sodium lauryl sulfate;magnesium lauryl sulfate; and the like. In one embodiment, the lubricantis stearic acid. In one embodiment, the lubricant is magnesium stearate.In one embodiment, the lubricant is magnesium stearate present in theamount of from about 0.5% to about 1.5% by weight of the formulation. Inone embodiment, the lubricant is magnesium stearate.

In one embodiment, the lubricant is present at an amount of about: 0.5%,0.75%, 1%, 1.25%, or 1.5 w/w. In another embodiment, the lubricant ispresent at an amount at an amount of about 0.5% w/w. In anotherembodiment, the lubricant is present at an amount at an amount of 0.5%w/w (±0.1%). In one embodiment, the lubricant is present at an amount of0.5% w/w (±0.2%). In such embodiments, the lubricant can be magnesiumstearate.

Binding agents or adhesives as used herein refer to an excipient whichimparts sufficient cohesion to the blend to allow for normal processingoperations such as sizing, lubrication, compression and packaging, butstill allow the formulation to disintegrate and the composition to beabsorbed upon ingestion. Exemplary binding agents and adhesives include,individually or in combination, acacia; tragacanth; glucose;polydextrose; starch including pregelatinized starch; gelatin; modifiedcelluloses including methylcellulose, carmellose sodium,hydroxypropylmethylcellulose (HPMC or hypromellose),hydroxypropylcellulose, hydroxyethylcellulose and ethylcellulose;dextrins including maltodextrin; zein; alginic acid and salts of alginicacid, for example sodium alginate; magnesium aluminum silicate;bentonite; polyethylene glycol (PEG); polyethylene oxide; guar gum;polysaccharide acids; and the like.

The binding agent(s) is present from about 2% to about 6%, by weight ofthe formulation. In one embodiment, the binding agent(s), is about 2%,3%, 4%, 5%, or 6 w/w. In another embodiment, the binder is present atabout 4% w/w of the formulation. In yet another embodiment, the binderis hypromellose.

Filler as used herein means an excipient that are used to dilute thecompound of interest prior to delivery. Fillers can also be used tostabilize compounds because they can provide a more stable environment.Salts dissolved in buffered solutions (which also can provide pH controlor maintenance) are utilized as diluents in the art, including, but notlimited to a phosphate buffered saline solution. Fillers increase bulkof the composition to facilitate compression or create sufficient bulkfor homogenous blend for capsule filling. Representative fillers includee.g., lactose, starch, mannitol, sorbitol, dextrose, microcrystallinecellulose such as Avicel®; dibasic calcium phosphate, dicalciumphosphate dihydrate; tricalcium phosphate, calcium phosphate; anhydrouslactose, spray-dried lactose; pregelatinized starch, compressible sugar,such as Di-Pac®(Amstar); hydroxypropylmethylcellulose,hydroxypropylmethylcellulose acetate stearate, sucrose-based diluents,confectioner's sugar; monobasic calcium sulfate monohydrate, calciumsulfate dihydrate; calcium lactate trihydrate, dextrates; hydrolyzedcereal solids, amylose; powdered cellulose, calcium carbonate; glycine,kaolin; mannitol, sodium chloride; inositol, bentonite, and the like.The filler(s) is present from about 6% to about 25%, by weight of theformulation. In one embodiment, the filler agent(s), is about 6%, 7%,8%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%,23%, 24%, or 25% w/w. In another embodiment, the composition comprisesabout 3.5% w/w or insoluble filler and about 2.5% w/w of soluble filler.In yet another embodiment, the insoluble filler is microcrystallinecellulose and the soluble filler is lactose.

As defined herein, where the mass of a compound is specified, forexample, “500 mg of compound (1),” that amount refers to the mass ofcompound (1) in its free base form.

The term “hemoglobin” as used herein refers to any hemoglobin protein,including normal hemoglobin (Hb) and sickle hemoglobin (HbS).

The term “sickle cell disease” (SCD) or “sicke cell diseases” (SCDs)refers to one or more diseases mediated by sickle hemoglobin (HbS) thatresults from a single point mutation in the hemoglobin (Hb). Sickle celldiseases includes sickle cell anemia, sickle-hemoglobin C disease(HbSC), sickle beta-plus-thalassaemia (HbS/β) and sicklebeta-zero-thalassaemia (HbS/β0).

“Substantially free” as used herein refers to ansolvate Form II ofCompound 1 associated with <10% or Form I and/or Form N, preferably <5%Form I and/or Form N; and most preferably it shall refer to <2% Form Iand/or Form N. Form I of Compound 1 is characterized by at least threeX-ray powder diffraction peaks (Cu Kα radiation) at 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andForm N of Compound 1 is characterized by at least three X-ray powderdiffraction peaks (Cu Kat radiation) at 11.65°, 11.85°, 12.08°, 16.70°,19.650 and 23.48°2θ (each ±0.2°2θ).

“Therapeutically effective amount” or “therapeutic amount” refers to anamount of a drug or an agent that when administered to a patientsuffering from a condition, will have the intended therapeutic effect,e.g., alleviation, amelioration, palliation or elimination of one ormore manifestations of the condition in the patient. The fulltherapeutic effect does not necessarily occur by administration of onedose, and may occur only after administration of a series of doses andcan be administered in one dose form or multiples thereof. For example,600 mg of the drug can be administered in a single 600 mg capsule or two300 mg capsules. Thus, a therapeutically effective amount may beadministered in one or more administrations. For example, and withoutlimitation, a therapeutically effective amount of an agent, in thecontext of treating disorders related to hemoglobin S, refers to anamount of the agent that alleviates, ameliorates, palliates, oreliminates one or more manifestations of the disorders related tohemoglobin S in the patient.

The term “pharmaceutically acceptable” refers to generally safe andnon-toxic for in vivo, preferably human, administration.

“Subject” or “patient” refers to human.

“Treatment”, “treating”, and “treat” are defined as acting upon adisease, disorder, or condition with an agent to reduce or amelioratethe harmful or any other undesired effects of the disease, disorder, orcondition and/or its symptoms. Treatment, as used herein, covers thetreatment of a human patient, and includes: (a) reducing the risk ofoccurrence of the condition in a patient determined to be predisposed tothe disease but not yet diagnosed as having the condition, (b) impedingthe development of the condition, and/or (c) relieving the condition,i.e., causing regression of the condition and/or relieving one or moresymptoms of the condition. For purposes of treatment of sickle celldisease, beneficial or desired clinical results include, but are notlimited to, multilineage hematologic improvement, decrease in the numberof required blood transfusions, decrease in infections, decreasedbleeding, and the like. For purposes of treatment of interstitialpulmonary fibrosis, beneficial or desired clinical results include, butare not limited to, reduction in hypoxia, reduction in fibrosis, and thelike.

6.2 Compounds and Uses

Compound 1 is2-hydroxy-6-((2-(1-isopropyl-1h-pyrazol-5-yl)pyridin-3-yl)methoxy)benzaldehyde,having the formula:

(hereinafter “Compound 1” or GBT440, where the terms are usedinterchangeably), or a tautomer thereof.

Compound 1 can be prepared according to the methods described in, forexample, International Publication Nos. WO 2015/031285 A1 (see, e.g.,pages 14-17) and WO 2015/120133 A1 (see, e.g., pages 32-35), thedisclosures of which are incorporated herein by reference in theirentireties.

The free base of Compound 1 can be obtained as one or more crystallineforms, such as those described in, for example, InternationalPublication Nos. WO 2015/031285 A1 (see, e.g., pages 19-24) and WO2015/120133 A1 (see, e.g., pages 3-9 and 51-54), including Form IIdescribed below.

Form II

In addition to the XRPD provided above, the crystalline Compound 1 ischaracterized by an endothermic peak at (97±2)° C. as measured bydifferential scanning calorimetry. In certain embodiments, thecrystalline Form II of the free base of crystalline Compound 1 ischaracterized by the substantial absence of thermal events attemperatures below the endothermic peak at (97±2)° C. as measured bydifferential scanning calorimetry. In certain embodiments, thecrystalline Form II of the free base of crystalline Compound 1 ischaracterized by an X-ray powder diffraction peak (Cu Kα radiation atone or more of 13.37°, 14.37°, 19.95° or 23.92°2θ. In certainembodiments, the crystalline ansolvate of the free base of crystallineCompound 1 is characterized by an X-ray powder diffraction peak (Cu Kαradiation at one or more of 13.37°, 14.37°, 19.95° or 23.92°2θ. Incertain embodiments, the crystalline Form IT of the free base ofcrystalline Compound 1 is characterized by an X-ray powder diffractionpattern (Cu Kα radiation) substantially similar to that of FIG. 1. Incertain embodiments, the crystalline ansolvate of the free base ofcrystalline Compound 1 is characterized by an X-ray powder diffractionpattern (Cu Kα radiation) substantially similar to that of FIG. 1.

In certain embodiments, the crystalline Form II of the free base ofcrystalline Compound 1 is characterized by at least one X-ray powderdiffraction peak (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ). In certain embodiments, the crystallineForm II of the free base of crystalline Compound 1 is characterized byat least two X-ray powder diffraction peaks (Cu Kα radiation) selectedfrom 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In certainembodiments, the crystalline Form II of the free base of crystallineCompound 1 is characterized by at least three X-ray powder diffractionpeaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ).

In certain embodiments, the crystalline ansolvate of the free base ofcrystalline Compound 1 is characterized by at least one X-ray powderdiffraction peak (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ). In certain embodiments, the crystallineansolvate of the free base of crystalline Compound 1 is characterized byat least two X-ray powder diffraction peaks (Cu Kα radiation) selectedfrom 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In certainembodiments, the crystalline ansolvate of the free base of crystallineCompound 1 is characterized by at least three X-ray powder diffractionpeaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ).

In certain embodiments, the crystalline ansolvate of the free base ofcrystalline Compound 1 is substantially free of Form I and/or Form N;wherein Form I is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andwherein Form N is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 11.65°, 11.85°,12.08°, 16.70°, 19.65° and 23.48°2θ (each ±0.2°2θ).

In certain embodiments, Form II is characterized by 1, 2, 3, 4, or morepeaks as shown in Table 1 below.

TABLE 1 Observed peaks for Form II, XRPD file 613881. Intensity °2θ dspace (Å) (%)  5.62 ± 0.20 15.735 ± 0.581 24 12.85 ± 0.20  6.888 ± 0.10822 12.97 ± 0.20  6.826 ± 0.106 21 13.37 ± 0.20  6.622 ± 0.100 100 14.37± 0.20  6.162 ± 0.087 56 15.31 ± 0.20  5.788 ± 0.076 21 16.09 ± 0.20 5.507 ± 0.069 23 16.45 ± 0.20  5.390 ± 0.066 69 16.75 ± 0.20  5.294 ±0.064 32 16.96 ± 0.20  5.227 ± 0.062 53 19.95 ± 0.20  4.450 ± 0.045 3920.22 ± 0.20  4.391 ± 0.043 20 23.18 ± 0.20  3.837 ± 0.033 38 23.92 ±0.20  3.721 ± 0.031 41 24.40 ± 0.20  3.648 ± 0.030 44 24.73 ± 0.20 3.600 ± 0.029 22 24.99 ± 0.20  3.564 ± 0.028 50 25.12 ± 0.20  3.545 ±0.028 28 25.39 ± 0.20  3.509 ± 0.027 51 25.70 ± 0.20  3.466 ± 0.027 2126.19 ± 0.20  3.403 ± 0.026 27 26.72 ± 0.20  3.336 ± 0.025 30 27.02 ±0.20  3.300 ± 0.024 25 27.34 ± 0.20  3.262 ± 0.024 23 28.44 ± 0.20 3.138 ± 0.022 20

In certain embodiments, Compound 1 is used in the treatment of sicklecell disease, as described herein. In certain embodiments, a polymorphof Compound 1, as described in any of the embodiments provided herein,is used in the treatment of sickle cell disease. In certain embodiments,a polymorph of the free base of crystalline Compound 1, as described inany of the embodiments provided herein, is used in the treatment ofsickle cell disease. In certain embodiments, the crystalline Form II ofthe free base of crystalline Compound 1, as described in any of theembodiments provided herein, is used in the treatment of sickle celldisease. In certain embodiments, the treatment is according to any ofthe pharmaceutical formulations, dosage forms, and/or dosage regimens asdescribed herein. In certain embodiments, such treatment comprisesadministering to a subject or preparing for administration to suchsubject,2-hydroxy-6-((2-(1-isopropyl-1H-pyrazol-5-yl)pyridin-3-yl)-methoxy)benzaldehyde,or a polymorph thereof, as described herein.

Accordingly, provided herein is a method for treating sickle celldisease in a patient comprising administering to the patient Compound 1.In certain embodiments, the compound is administered in a dose of fromabout 500 mg/day to about 1500 mg/day. In certain embodiments, thecompound is administered in a dose of from about 600 mg/day to about 900mg/day. In certain embodiments, the compound is administered in a doseof about 600 mg/day. In certain embodiments, the compound isadministered in a dose of about 900 mg/day, or about 1200 mg/day, orabout 1500 mg/day. In certain embodiments, the compound is administeredin a dose of 600 mg/day. In certain embodiments, the compound isadministered in a dose of 900 mg/day, 1200 mg/day or 1500 mg/day. Incertain embodiments, the compound is administered once daily. In certainembodiments, the compound is a crystalline ansolvate form of Compound 1as described in any of the embodiments provided herein.

Accordingly, also provided herein is Compound 1 for use in the treatmentof sickle cell disease. In certain embodiments, about 900 mg/day toabout 1500 mg/day of the compound is used for treatment. In certainembodiments, about 900 mg/day, about 1200 mg/day, or about 1500 mg/dayof the compound is used for treatment. In certain embodiments, 900mg/day, 1200 mg/day, or 1500 mg/day of the compound is used fortreatment. In certain embodiments, the compound is used for treatment asa single dose. In certain embodiments, the compound is a crystallineansolvate form of Compound 1 as described in any of the embodimentsprovided herein. In certain embodiments, the compound is prepared foruse as a medicament, for example, a pharmaceutical formulation or dosageform, as described herein.

6.3 Pharmaceutical Formulations and Dosage Forms

In another aspect, Compound 1 is administered in a pharmaceuticalformulation. Accordingly, provided herein are pharmaceuticalformulations comprising a pharmaceutically acceptable excipient and acompound disclosed herein. In certain embodiments, the pharmaceuticalformations comprise the crystalline free base ansolvate of Compound 1,including, for example, crystalline Form II. Suitable formulations arethose described in, for example, International Publication No. WO WO2015/031284 A1 (see, e.g., pages 18-21 and 28-29), the disclosure ofwhich is incorporated herein by reference in its entirety.

Such formulations can be prepared for different routes ofadministration.

Although formulations suitable for oral delivery will probably be usedmost frequently, other routes that may be used include intravenous,intramuscular, intraperitoneal, intracutaneous, and subcutaneous routes.Suitable dosage forms for administering any of the compounds describedherein include tablets, capsules, pills, powders, parenterals, and oralliquids, including suspensions, solutions and emulsions. Sustainedrelease dosage forms may also be used. All dosage forms may be preparedusing methods that are standard in the art (see, e.g., Remington'sPharmaceutical Sciences, 16th ed., A. Oslo editor, Easton Pa. 1980).Because of their ease of administration, tablets and capsules representthe most advantageous oral dosage unit forms.

Pharmaceutically acceptable excipients are generally non-toxic, aidadministration, and do not adversely affect the therapeutic benefit ofCompound 1. Such excipients may be any solid, liquid, semi-solid or, inthe case of an aerosol composition, gaseous excipient that is generallyavailable to one of skill in the art. The pharmaceutical compositionsdisclosed herein are prepared by conventional means using methods knownin the art.

The formulations disclosed herein may be used in conjunction with any ofthe vehicles and excipients commonly employed in pharmaceuticalpreparations, e.g., talc, gum arabic, lactose, starch, magnesiumstearate, cocoa butter, aqueous or non-aqueous solvents, oils, paraffinderivatives, glycols, etc. Coloring and flavoring agents may also beadded to preparations, particularly to those for oral administration.Solutions can be prepared using water or physiologically compatibleorganic solvents such as ethanol, 1,2-propylene glycol, polyglycols,dimethylsulfoxide, fatty alcohols, triglycerides, partial esters ofglycerin and the like.

Solid pharmaceutical excipients include starch, cellulose, hydroxypropylcellulose, talc, glucose, lactose, sucrose, gelatin, malt, rice, flour,chalk, silica gel, magnesium stearate, sodium stearate, glycerolmonostearate, sodium chloride, dried skim milk and the like. Liquid andsemisolid excipients may be selected from glycerol, propylene glycol,water, ethanol and various oils, including those of petroleum, animal,vegetable or synthetic origin, e.g., peanut oil, soybean oil, mineraloil, sesame oil, etc. In certain embodiments, the compositions providedherein comprises one or more of α-tocopherol, gum arabic, and/orhydroxypropyl cellulose.

The amounts of active ingredients in a dosage form may differ dependingon factors such as, but not limited to, the route by which it is to beadministered to patients. In certain embodiments, the dosage formsprovided herein comprise Compound 1 in an amount of about 10, about 20,about 30, about 40, about 50, about 100, about 150, about 200, about250, about 300, about 400, or about 500 mg. In certain embodiments, thedosage forms provided herein comprise Compound 1 in an amount of about:500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1050, 1100,1150, 1200, 1250, 1300, 1350, 1400, 1450, or 1500 mg. In certainembodiments, the dosage forms provided herein comprise Compound 1 in anamount of about: 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450,or 1500 mg. In certain embodiments, the dosage forms provided hereincomprise Compound 1 in an amount of about 50, about 100, or about 300mg. In certain embodiments, the dosage forms provided herein compriseCompound 1 in an amount of about 300, about 600, about 900, about 1200,or about 1500 mg. In certain embodiments, the dosage forms providedherein comprise Compound 1 in an amount of 300 mg±10%. In certainembodiments, the dosage forms provided herein comprise Compound 1 in anamount of 300 mg±5%.

In one embodiment, provided is a capsule dosage form described in theSummary above (and embodiments thereof). The formulation in the capsuleis prepared by wet granulation process as described below.

1. Dispensing

All the ingredients except the lubricant is screened through a 20-meshscreen to remove any agglomerates. The lubricant is screened through a40-mesh screen.

2. High Shear Wet Granulation (HSWG) and Fluid Bed Drying

All the ingredients screened in the dispensing step except for thelubricant are added in a predefined order to the wet granulation bowl.The ingredients are mixed in the granulation bowl using the impelloronly for a predetermined time to form a homogenous dry mixture. To thedry mix, water is used as a binding solution at a predetermined rate andamount while mixing using a high shear force with impellor and chopperat predetermined speeds. After adding the required amount of water, thewet granulation in kneaded or wet massed using both the impellor andchopper at predetermined speed and time. The wet granulation obtained isthen transferred to the fluid bed dryer for drying. The granulation isdried until the desired dryness level is achieved measured by loss ondrying (LOD)

3. Co-Milling or Sizing and Blending

The dried granulation from the HSWG and FBD step is then sized using aco-mill with a predetermined screen size and speed. A co-mill is used asa sizing step to ensure deagglomeration of large granule agglomeratesand help achieve a uniform particle size distribution. The driedgranules are then blended for a predetermined time in a V-blender alongwith the lubricant until a homogenous uniform blend is obtained. Thefinal blend is then transferred to the encapsulation process.

4. Encapsulation, Packaging and Labeling

The final granulation blend is filled into capsules using either asemi-automatic/manual encapsulator or an automatic encapsulatordepending on the scale and availability. A target weight of 350 mg ofthe granulation (containing 300 mg of API) is filled into each emptycapsule to make 300 mg strength capsules. Filled capsules are polishedfollowed by weight check and visual inspection for appearance to removeany defective capsules. Capsules are then packaged into 100 cchigh-density polyethylene (HDPE) bottles at 30 capsules per bottle. TheHDPE bottles are closed with child-resistant polypropylene (PP) screwcaps with liner. Appropriate labels are applied over the HDPE bottles asper the regional regulations.

6.4 Capsule Dosage Forms

In certain embodiments, the capsule dosage form comprises:

(i) from about 65% to about 93% w/w of Compound 1 or a polymorphthereof; and

(ii) from about 2% to about 10% w/w a binder;

wherein w/w is relative to the total weight of the formulation(excluding the weight of the capsule). With regards to the capsuleformulation; “about” means ±10% of a given range or value.

In certain embodiments, the capsule dosage form further comprises fromabout 2% to about 10% a disintegrant.

In certain embodiments, the capsule dosage form further comprises fromabout 2% to about 10% a disintegrant and about 2% to 35% a filler.

In certain embodiments, the capsule dosage form comprises:

(i) from about 65% to about 86% w/w of Compound 1 or a polymorphthereof;

(ii) from about 2% to about 6% w/w a binder;

(iii) from about 6% to about 25% w/w a filler;

(iv) from about 2% to 6% w/w a disintegrant; and

(iv) from about 0.5% to about 1.5% w/w a lubricant;

wherein w/w is relative to the total weight of the formulation(excluding the weight of the capsule). With regards to the capsuleformulation; “about” means ±10% of a given range or value.

In certain embodiments, the capsule dosage form comprises:

(i) from about 65% to about 86% w/w of Compound 1 or a polymorphthereof;

(ii) from about 2% to about 6% w/w a binder;

(iii) from about 3.5% to about 25% w/w an insoluble filler or 2.5% to25% w/w of soluble filler or 2.5% to 25% of a combination of soluble orinsoluble filler;

(iv) from about 2% to 6% w/w a disintegrant; and

(iv) from about 0.5% to about 1.5% w/w a lubricant.

In certain embodiments, the capsule dosage form comprises:

(i) about 86% w/w of Compound 1 or a polymorph thereof;

(ii) about 4% w/w a binder;

(iii) about 3.5% w/w an insoluble filler and 2.5% w/w of soluble filler;

(iv) about 3.5% w/w a disintegrant; and

(iv) about 0.5% w/w a lubricant.

In certain embodiments, the capsule dosage form comprises:

(i) 85.71% w/w of Compound 1 or a polymorph thereof;

(ii) 4% w/w a binder;

(iii) 3.64% w/w an insoluble filler and 2.65% w/w of soluble filler;

(iv) 2.65% w/w a disintegrant; and

(iv) 0.5% w/w a lubricant.

In certain embodiments:

Compound 1 is Form II substantially free of Form I and/or N;

the binder is hypromellose;

the insoluble filler is microcrystalline cellulose

the soluble filler is lactose monohydrate;

the disintregrant is croscarmellose sodium; and

the lubricant is magnesium stearate.

In certain embodiments, the capsule contains 300 mg of Compound 1 FormIT substantially free of Form I and/or N.

In certain embodiments, Compound 1 is a crystalline ansolvate form. Inone embodiment, the crystalline ansolvate is Form II characterized by atleast one X-ray powder diffraction peak (Cu Kα radiation) selected from13.37°, 14.37°, 19.95° and 23.92°2θ (each f0.2°2θ). In one embodiment,the crystalline ansolvate is characterized by at least one X-ray powderdiffraction peak (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ). In another embodiment, Form II ischaracterized by at least two X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In another embodiment, the crystalline ansolvate ischaracterized by at least two X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ). In yet another embodiment, Form II is characterized by atleast three X-ray powder diffraction peaks (Cu Kα radiation) selectedfrom 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yet anotherembodiment, the crystalline ansolvate is characterized by at least threeX-ray powder diffraction peaks (Cu Kα radiation) selected from 13.37°,14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yet another embodiment,Form II is characterized by X-ray powder diffraction peaks (Cu Kαradiation) of 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). In yetanother embodiment, the crystalline ansolvate is characterized by X-raypowder diffraction peaks (Cu Kα radiation) of 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ). In yet another, Form II is characterized by anX-ray powder diffraction pattern (Cu Kα radiation) substantially similarto that of FIG. 1. In yet another, the crystalline ansolvate ischaracterized by an X-ray powder diffraction pattern (Cu Kα radiation)substantially similar to that of FIG. 1.

In certain embodiments, the capsule contains 300 mg±5% of Compound 1,wherein compound 1 is a crystalline ansolvate form that is characterizedby at least two X-ray powder diffraction peaks (Cu Ku radiation)selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ);wherein the crystalline ansolvate form is substantially free of Form Iand/or N; wherein Form I is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andwherein Form N is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 11.65°, 11.85°,12.08°, 16.70°, 19.65° and 23.48°2θ (each ±0.2°2θ).

6.5 Dosages

The dose of the compounds disclosed herein to be administered to apatient can be subject to the judgment of a health-care practitioner.Doses of the compounds disclosed herein vary depending on factors suchas: specific indication to be treated, prevented, or managed; age andcondition of a patient; and amount of second active agent used, if any.

In certain embodiments, the compound (e.g., Compound 1) is administeredin a dose of from about 500 mg/day to about 1500 mg/day. In oneembodiment the compound is administered in a dose of about 1100, about1200, about 1300, about 1400, or about 1500 mg/day. In certainembodiments, the compound is administered in a dose of about isadministered in a dose of about 500, about 550, about 600, about 650,about 700, about 750, about 800, about 850, about 900, about 950, about1000, about 1050, about 1100, about 1150, about 1200, about 1250, about1300, about 1350, about 1400, about 1450, or about 1500 mg/day. Incertain embodiments, the compound is administered in a dose of about1050, about 1100, about 1150, about 1200, about 1250, about 1300, about1350, about 1400, about 1450, or about 1500 mg/day. In certainembodiments, the compound is administered in a dose of about 500, about550, about 600, about 650, about 700, about 750, about 800, about 850,about 900, about 950, or about 1000 mg/day. In certain embodiments, thecompound is administered in a dose of about 600, about 650, about 700,about 750, about 800, about 850, or about 900 mg/day. In certainembodiments, the compound is administered in a dose of from about 500mg/day to about 900 mg/day. In certain embodiments, the compound isadministered in a dose of from about 600 mg/day to about 900 mg/day. Incertain embodiments, the compound is administered in a dose of about 700mg/day. In certain embodiments, the compound is administered in a doseof about 600 mg/day. In certain embodiments, the compound isadministered in a dose of about 900 mg/day. In certain embodiments, thecompound is administered in a dose of about 1200 mg/day. In certainembodiments, the compound is administered in a dose of about 1500mg/day.

In certain embodiments, the compound (e.g., Compound 1) is administeredas mg/Kg body weight of the patient, for example, from about 5 to about50 mg/Kg body weight of the patient being treated/day, from about 10 toabout 40 mg/Kg/day, from about 15 to about 30 mg/Kg/day, from about 15to about 25 mg/Kg/day, about 5 to about 10 mg/Kg/day, about 10 to about15 mg/Kg/day, about 15 to about 20 mg/Kg/day, about 20 to about 25mg/Kg/day, about 25 to about 30 mg/Kg/day, about 30 to about 40mg/Kg/day, or about 40 to about 50 mg/Kg/day.

The dose may be administered as a single bolus, or in one, two, three,four or more subdoses at appropriate intervals throughout the day. Forexample, if the dose to be administered is 900 or 1500 mg/day, theentire 900 or 1500 mg, respectively, may be administered at the sametime. Alternatively, the 900 mg dose may be administered as, forexample, three separate subdoses of 300 mg, where the first subdose isadministered in the morning, the second subdose is administered in theafternoon of the same day, and the third subdose is administered in theevening of the same day, such that the cumulative amount administeredfor the day is 900 mg.

7. EXAMPLES

Certain embodiments disclosed herein are illustrated by the followingnon-limiting examples.

7.1 Example 1

The following example presents a Phase I randomised, placebo-controlled,double-blind, single and multiple ascending dose study of thetolerability and pharmacokinetics of Compound 1 (GBT440) in healthysubjects and patients with Sickle Cell Disease.

Objectives:

Primary Outcome Measures:

-   -   Safety, as assessed by frequency and severity of adverse events        (AEs), and changes in vital signs, 12-lead electrocardiograms        (ECGs), and laboratory assessments as compared to baseline [Time        Frame: 30 days]

Secondary Outcome Measures:

-   -   Blood and plasma area under the concentration time curve (AUC)        of GBT440 [Time Frame: 30 days]    -   Blood and plasma maximum concentration (Cmax) of GBT440 [Time        Frame: 30 days]    -   Blood and plasma time to maximum concentration (Tmax) of GBT440        [Time Frame: 30 days]    -   Percentage of hemoglobin occupied or modified by GBT440 [Time        Frame: 30 days]    -   Change from baseline in heart rate and pulse oximetry following        exercise testing in healthy volunteers [Time Frame: 30 days]

Other Outcome Measures:

-   -   Percentage of sickled cells under ex vivo conditions [Time        Frame: 30 days]    -   Effect of GBT440 on hemolysis as measured by LDH, direct        bilirubin, hemoglobin, and reticulocyte count [Time Frame: 30        days]    -   Change from baseline in pain as measured by visual analog scale        [Time Frame: 30 days]    -   Change from baseline in fatigue as measured by questionnaire        [Time Frame: 30 days]    -   Exercise capacity as measured by 6-minute walk test [Time Frame:        30 days]

Methodology:

Experimental: GBT440

-   -   Subjects randomized 6:2 to receive daily oral dosing of GBT440        or placebo for 1 day (single dose) and up to 28 days (multiple        dose)

Placebo Comparator: Placebo

-   -   Subjects randomized 6:2 to receive daily oral dosing of GBT440        or placebo for 1 day (single dose) and up to 28 days (multiple        dose)

Number of Subjects: 128

Criteria:

Inclusion Criteria:

-   -   Healthy male or female of non-child bearing potential; 18 to 55        years old; are non-smokers and have not used nicotine products        within 3 months prior to screening.    -   Male or female, 18 to 60 years old, with sickle cell disease        (hemoglobin SS) not requiring chronic blood transfusion therapy;        without hospitalization in 30 days before screening or receiving        blood transfusion within 30 days before screening; subjects are        allowed concomitant use of hydroxyurea if the dose has been        stable for the 3 months prior to screening.

Exclusion Criteria:

-   -   Subjects who have a clinically relevant history or presence of        respiratory, gastrointestinal, renal, hepatic, haematological,        lymphatic, neurological, cardiovascular, psychiatric,        musculoskeletal, genitourinary, immunological, dermatological,        connective tissue diseases or disorders.    -   Subjects who consume more than 14 (female subjects) or 21 (male        subjects) units of alcohol a week.    -   Subjects who have used any investigational product in any        clinical trial within 90 days of admission or who are in        extended follow-up.    -   Healthy subjects who have used prescription drugs within 4 weeks        of first dosing or have used over the counter medication        excluding routine vitamins within 7 days of first dosing.    -   Subjects with sickle cell disease who smoke >10 cigarettes per        day; have hemoglobin level <6 mg/dL or >10 mg/dL at screening;        have aspartate aminotransferase (AST), alanine aminotransferase        (ALT), or alkaline phosphatase (ALK) >3× upper limit of normal        reference range (ULN) at screening; have moderate or severe        renal dysfunction

Test Product, Dose and Route of Administration:

Compound 1 oral capsules at 2 strengths (50 and 100 mg)

Doses: 300, 500, 600, 700, 900, or 1000 mg/day

Alternatively, the following Doses may also be used: 900, 1200, or 1500mg/day.

7.2 Example 2

The following example presents pharmacokinetic results from the study asdescribed in Example 1.

Analysis of whole blood was performed as follows. 50 μL of diluted wholeblood was mixed with 20 μL of GBT1592 (GBT440-D7) solution inacetonitrile. 0.3 mL of 0.1M citrate buffer solution (pH 3) was added tothe sample, and the sample mixed briefly by vortexing, followed bysonication for 10 minutes. 2.0 mL methyl tert butyl ether (MTBE) wasadded to the sample, and the sample was capped, and mixed thoroughly byvortexing at high speed for 20 minutes. The sample was then centrifugedat 3300 rpm at room temperature for 10 minutes. 0.2 mL of the clearorganic layer of the centrifuged sample was then transferred to a clean96-well 2-mL plate, and the solvent was evapored to dryness. The driedextract was reconstituted in 0.2 mL of a mixture ofacetonitrile/methanol/water/DMSO (225:25.0:250:50.0) and mixedthoroughly. The resultant reconstituted extract was analyzed by liquidchromatography mass spectrometry (LCMS).

For the LCMS, a Sciex API 4000 LC-MS-MS was equipped with an HPLCcolumn. The peak area of the m/z 338.1→158.1 GBT440 product ion wasmeasured against the peak area of the m/z 345.2→159.1 GBT1592(GBT440-D7) internal standard product ion.

The whole blood samples, obtained as described above, were analyzed forpharmacokinetic parameters and RBC:Plasma ratios, as follows.

Terminal half-life and other pharmacokinetic parameters were calculatedusing Phoenix WinNonlin software. Apparent terminal half-life (t_(1/2))values were calculated as ln(2)/k, where k is the terminal eliminationrate constant which is obtained by performing a linear regression on theterminal phase of a plot of the natural logarithm (ln) of concentrationversus time.

RBC:Plasma ratio was calculated using the equation below.

$\begin{matrix}{\frac{RBC}{PL} - \frac{\frac{BL}{PL} - \left( {1 - {Hct}} \right)}{Hct}} & {{Equation}\mspace{14mu} 1}\end{matrix}$

In Equation 1, RBC is the concentration of GBT440 in the red bloodcells; PL is the concentration of GBT440 in plasma obtained by analysisof plasma sample; BL is the concentration of GBT440 in whole bloodobtained by analysis of whole blood sample; and Hct is the hematocritvalue.

FIG. 2 illustrates representative whole blood concentrations at steadystate for two doses (500 mg, 700 mg) of Compound 1 (GBT440).

A dose proportional increase in GBT440 was observed following single andmultiple dosing. From these pharmacokinetic studies, the half-life ofGBT440 in whole blood was determined to be approximately 3 days inhealthy subjects, and 1.6 days in SCD subjects. In the tested subjects,the GBT440 RBC:plasma ratio was observed to be approximately 75:1. Thesepharmacokinetic results support once daily dosing.

7.3 Example 3

The following example presents hemoglobin oxygen equilibration results(e.g., oxygen equilibration curves) following dosing with Compound 1(GBT440), from the study as described in Example 1.

Whole blood hemoximetry was used to measure oxygen equilibration. Bloodfrom healthy volunteers and sickle cell disease (SCD) patients was drawninto 1.8 mL sodium citrate tubes. These samples were stored overnight at4° C. prior to hemoximetry measurements. Based upon the hematocrit ofthe blood, either 50 μL or 100 μL of blood was diluted into 5 mL of 37°C. TES buffer (30 mM TES, 130 mM NaCl, 5 mM KCl, pH 7.4 at 25° C.).Diluted sample were loaded into TCS Hemox Analyzer cuvettes andoxygenated for twenty minutes using compressed air. After oxygenation,the samples were deoxygenated using nitrogen gas until the pO₂ reached1.6 millimeters of mercury (mm Hg). Data during this deoxygenation stepwas collected into Oxygen Equilibrium Curve (OEC) files using the TCSHemox Analytical Software (HAS). OEC files were then analyzed to obtainthe p50 (the partial pressure of oxygen at which 50% of hemoglobin in asample is saturated with O₂) and the p20 (the partial pressure of oxygenat which 20% of hemoglobin in a sample is saturated with O₂). Delta p20values (p20_(predose)−p20_(sample time)) were then used to calculate the% Hb Modification.

FIG. 3 illustrates representative oxygen equilibrium curves for twodoses (500 mg, 700 mg) of Compound 1 (GBT440), with comparison toplacebo. As shown in this figure, administration of Compound 1 resultsin a left shift of the oxygen equilibrium curve: SCD subjects are rightshifted; p50 shifts to normal range. As also shown in this figure,hemoglobin modification is proportional to dose.

7.4 Example 4

The following example presents results showing a change in hemoglobinover time following dosing with Compound 1 (GBT440), from the study asdescribed in Example 1.

FIG. 4 illustrates the change in hemoglobin (g/dL) over time for twodoses (500 mg, 700 mg) of Compound 1 (GBT440), with comparison toplacebo. As shown in the figure, GBT 440 treatment led to a rapid andprogressive rise in hemoglobin levels. The decline in later time pointsmay be related to removal of dense cells and not related to return ofhemolysis. The higher GBT440 dose level (700 mg) showed a trend for abetter response compared to 500 mg. These results show that a reductionin hemolysis increases hemoglobin levels.

7.5 Example 5

The following example presents results showing a change in reticulocytes(e.g., percent change in reticulocytes) over time following dosing withCompound 1 (GBT440), from the study as described in Example 1.

FIG. 5 illustrates the percent (%) change in reticulocytes over time fortwo doses (500 mg, 700 mg) of Compound 1 (GBT440), with comparison toplacebo. As shown in the figure, GBT 440 treatment led to a profounddecline in reticulocytes, which is consistent with a reduction inhemolysis. The reduction in reticulocyte counts suggests improvement ofred blood cell life span.

7.6 Example 6

The following example presents results showing a change in circulatingsickle cells (e.g., percent change in circulating sickle cells) overtime following dosing with Compound 1 (GBT440), from the study asdescribed in Example 1.

FIG. 6 illustrates percent (%) sickle cells over time for two doses (500mg, 700 mg) of Compound 1 (GBT440), with comparison to placebo. As shownin the figure, baseline sickle cell counts were variable (1.1 to 19.4%).As also shown in the figure, GBT440 treatment reduced sickle cells inthe peripheral blood which was sustained during the 28 day dosingperiod. These results show that a reduction in hemolysis increaseshemoglobin levels.

This example also provides results showing a change in circulatingsickle cells (e.g., percent change in circulating sickle cells) overtime following dosing with Compound 1 (GBT440), from the study asdescribed in Example 1.

To quantify irreversibly sickled cells (ISCs), six different fields wererandomly selected and imaged at 40× magnification per slide. Each fieldcontained 100 to 300 cells and >500 cells (in 3 or more fields) werecounted per blood smear slide. Cells that were classically sickled shapeor appeared linear (with length equal to or more than 3× the width) withirregular or pointed edges were counted as sickled. Elliptical red bloodcells (also appearing linear but with length approximately twice thewidth) with smooth rounded edges were counted as normal. In general,isolated non-discoid cells with spiky turns were counted as sickled.Cells packed in a group that appeared non-discoid because of the packingwere not counted as sickled since they demonstrate deformability bychanging shape to accommodate the surrounding cells.

Morphological criteria for sickle cells included the followingcategories: (1) non-discoid irregular shaped cells with irregular orpointed edges; (2) elliptocytes with length more than twice the widthand with irregular or pointed edges; and (3) irregular shapedelliptocytes.

FIGS. 7A and 7B provide representative images from a subject treatedwith 700 mg of Compound 1 (GBT440), over a period of one day as shown inFIG. 7A; and twenty-eight (28) days as shown in FIG. 7B. As shown in thefigure, there is a marked reduction in sickle cells in peripheral bloodsmears.

7.7 Example 7

The following example presents results showing a change in reticulocytesat day 28, as a function of whole blood concentration of Compound 1(GBT440), from the study as described in Example 1.

The strongest correlation between exposure and hematologic effect wasobserved with changes in reticulocyte counts (considered to be bestbiomarker for RBC survival).

FIG. 8 illustrates the percent (%) change in reticulocytes to day 28versus whole blood concentration of Compound 1 (GBT440) (PK data from500 and 700 mg dose levels; R²˜0.56). As shown in the figure, higherGBT440 exposures resulted in more profound reduction in reticulocytecounts.

The results provided in the above Examples 1-7 for Compound 1 (GBT440)demonstrate favorable pharmacokinetic data (e.g., long terminal t %), exvivo anti-sickling activity, ability to increase hemoglobin levels, andability to reduce reticulocyte counts. Further, the results provided inthese Examples demonstrate that GBT440 whole blood concentrations weremuch higher than plasma concentrations (RBC:plasma ratio ˜75:1),consistent with a high affinity and specificity of GBT440 forhemoglobin. These results supports the potential Compound 1 (GBT440) tobe a beneficial therapeutic agent, suitable for once daily dosing at thedisclosed doses, for the treatment of SCD.

7.8 Example 8

The following example presents response analysis of Compound 1 (GBT440)based on PK/PD modeling and hemolysis measures.

A PK/PD model was developed using PK and PD data from subjects with SCD,corresponding to Cohorts 11 (700 mg QD×28 days), 12 (500 mg QD×28 days)and 14 (500 mg BID×28 days) who participated in the study described inExample 1 above. The PK/PD model was developed to characterize therelationship between Compound 1 (GBT440) exposures, placebo andhemolysis measures (e.g., reticulocyte count, hemoglobin, unconjugatedbilirubin and LDH). The drug effect was characterized using an indirectresponse model of drug/dose or concentration-dependent inhibition (e.g.,bilirubin, reticulocytes, and LDH) or drug/dose orconcentration-dependent stimulation (e.g., hemoglobin). Linear andnon-linear models (maximal effect, e.g., E_(max) model and sigmoidalE_(max) model) were explored while the PK part of the model was keptfixed (e.g., sequential analysis). The PK/PD model used for hemolyticmeasures is shown in the equation below.

$\begin{matrix}{\frac{d{A(1)}}{dt} = {{k_{in} \times \left( {1 - {{Sl} \times {WBC}_{GBT440}}} \right)} - {k_{out} \times {A(1)}}}} & {{Equation}\mspace{14mu} 2}\end{matrix}$

where

A(1)_(initial)=Base  (Equation 3)

In Equation 2, A(1) represents the amount of biomarker of interest; Slrepresents the slope of the drug effect; WBC_(GBT440) is the whole bloodconcentration of GBT440; and k_(in) and k_(out) are the production rateand the disappearance rate constant, respectively, of each biomarker.

The ratio of k_(in) and k_(out) represents the baseline of the biomarkerat steady state, as shown in the equation below.

$\begin{matrix}{{Base} = \frac{k_{in}}{k_{out}}} & {{Equation}\mspace{14mu} 4}\end{matrix}$

The final PK/PD relationship for the hemolysis markers was bestdescribed with an indirect response model where drug-related efficacywas driven by Compound 1 (GBT440) whole blood pharmacokinetics. Linearexposure response models were sufficient to characterize the data.

Based on this modeling, it was determined that the PD effects for thehemolysis measures (e.g., bilirubin, reticulocyte count, LDH andhemoglobin) are PK driven. FIGS. 9A-9B illustrate the linearrelationship between Compound 1 whole blood concentrations and effect onhemolytic measures: FIG. 9A shows percent (%) change in absolutereticulocytes; FIG. 9B shows percent (%) change in unconjugatedbilirubin; FIG. 9C shows percent (%) change in LDH; and FIG. 9D showspercent (%) change in hemoglobin. In the figure, the dashed linerepresents predicted change for a typical patient, the grey shaded arearepresents 95% CI (uncertainty in relationship), and the dotted linesrepresent 2.5^(th) and 97.5^(th) percentiles of the 600 mg and the 900mg dose. The drug-related efficacy is a function of bloodpharmacokinetics and the PD effects for the hemolysis measures disappearafter dosing is stopped. A linear concentration-effect relationship wasobserved over the range of doses evaluated (500 mg to 1000 mg).

7.9 Example 9

The following example presents Hb occupancy analysis of Compound 1(GBT440) based on population PK modeling. The following examples alsopresents simulated SCD measures outcomes.

Hb Modification (% Occupancy):

A population PK model was developed for Compound 1 (GBT440) based ondata from healthy subjects and patients participating in the study asdescribed in Example 1. The population PK model was developed todetermine which doses would achieve Hb occupancy from 20% to 30%, whichis the target range for therapeutic efficacy with Compound 1. The targetrange of 20% to 30% Hb modification is supported by treatment responsedata from the study. Participants who achieved >20% Hb occupancy showedan improved hematologic response compared to those who did not whoachieve >20% Hb occupancy. Population PK models were developed forCompound 1 measured in plasma and in whole blood. Separate models weredeveloped for patients and healthy subjects, as these populationsappeared to show substantial differences in Compound 1 PK, due to thenature of SCD.

The percent Hb modification (% occupancy) was calculated according toEquations 5 and 6 below, where whole blood and plasma concentrationswere derived from the population PK model, and hematocrit values (Hct)values were uniformly sampled from the range available in the database.A constant of 0.3374 was used in Equation 5 to convert RBC concentrationfrom μg/mL into μM. In Equation 6, % occupancy was defined as theconcentrations of Compound 1 in RBC (in μM) divided by the concentrationof Hb in RBC (5000 μM). The models were used to evaluate the potentialof several Compound 1 doses (e.g., 900 mg, 1200 and 1500 mg) to achievethe occupancy target of 20% to 30%.

$\begin{matrix}{{RBC}_{conc} = {\frac{{Blood}_{conc} - \left\lbrack {\left( {1 - {Hct}} \right) \times {Plasma}_{conc}} \right\rbrack}{Hct} \times \frac{1}{0.3374}}} & {{Equation}\mspace{14mu} 5} \\{\mspace{79mu}{{{Calculated}\mspace{14mu}\%\mspace{14mu}{Occupancy}} = \frac{{RBC}_{conc} \times 100}{1000 \times 5}}} & {{Equation}\mspace{14mu} 6}\end{matrix}$

TABLE 2 Hb Occupancy Target for Compound 1 at doses of 900 mg and 1500mg Dose of GBT440 Estimated Hb Occupancy 900 mg 1500 mg Median %occupancy 16 (7-31) 26 (12-52) based on C_(min) (2.5^(th) to 97.5^(th)percentiles) % Subjects with >20% 24.6% 75.5% occupancy based on C_(min)Values based on modeling of PK/PD data derived from the study asdescribed in Example 1 and further simulations of such data. Linearpharmacokinetics has been assumed for simulations of 1500 mg dose.

Additionally, determination of the estimated change from baseline inhemolysis measures for 900 mg and 1500 mg doses based on simulations(see Table 3 below) showed improvement over those observed in Cohorts11, 12 and 14 (see Table 4 below).

TABLE 3 Simulated SCD Measures Outcomes (% Change from Baseline) forCompound 1 at doses of 900 mg and 1500 mg Compound 1 Doses HemolysisMeasure 900 mg 1500 mg Bilirubin (%) −47(33-62)^(a) −66 (51-78)^(a)Reticulocytes (%) −54(28 − 78)^(a) −84 (61-94)^(a) LDH (%) −30(13-56)^(a) −64 (37-84)^(a) Hemoglobin (%) 11.9 (6.7 − 21.1)^(b) ^(d)Hemoglobin (change 1.06 (0.60 − 1.9)^(c) ^(d) from baseline) Valuesrepresent median (25^(th) to 97.5^(th) percentiles) ^(a)Based on E_(max)model. Note: An E_(max) model was used to fit the hemolysis measuresdata. The E_(max) model provided a similar fit to the bilirubin,reticulocytes and LDH data as the linear model, however it requiredE_(max) value to be fixed to 100%, (these measures are decreasing overtime). Since hemoglobin increases over time, the E_(max) model was lessrobust than the linear model (Δ OFV >25). Therefore predictions were notattempted for hemoglobin outside of the observed dose range (e.g. >1000mg). ^(b)Based on linear model ^(c)Based on a baseline Hb of 9 g/dL^(d)For hemoglobin measurements, the E_(max) model resulted in a lessreliable fit, with more uncertain estimates of E_(max) and EC₅₀(RSE >100%), and therefore was not used to make predictions for the 1500mg dose. The linear model was satisfactory describing the data in theobserved dose range, however the linear model should not be used toextrapolate to higher doses, however, it can be assumed that treatmentresponse of the higher dose (1500 mg) will be at least equal or highercompared to the lower dose (900 mg).

TABLE 4 Change from Baseline to Day 28 in Response Parameters inSubjects with SCD (Cohorts 11, 12, and 14) Change from Baseline to Day28 Median (25^(th), 75^(th) percentile) GBT440 GBT440 GBT440 500 mg 700mg 1000 mg^(a) Placebo (Cohort 12) (Cohort 11) (Cohort 14) (Pooled)Parameter n = 10 n = 12 n = 5 n = 12 Unconjugated −30.6 −42.6 −56.3 2.0bilirubin (%) (−48.9, (−44.3, (−57.8, (−24.6, −15.4) −23.8) −47.1) 9.9)Reticulocytes −31.2 −37.0 −49.9 9.0 (%) (−48.9, (−52.6, (−64.3, (1.7,−20.8) −4.5) −34.4) 13.8) Hemoglobin 0.4 0.7 0.0 −0.1 (g/dL) (0.1, (0.5,(−0.4, (−0.3, 0.7) 1.0) 0.3) 0.4) Lactate −19.8 −11.9 −12.4 −4.8dehydrogenase (%) (−39.0, (−30.1, (−20.2, (−13.1, 6.2) −5.7) −12.2)−2.3) Irreversibly −56.4 −45.9 −45.7 8.4 sickled cells (%) (−70.2,(−93.0, (−57.9, (−11.9, −26.2) −6.0) 5.9) 16.8) ^(a)500 mg twice dailySource: Listing lb_2 for hemoglobin and Listing lb2_2 for LDH,bilirubin, and reticulocytes for 28-day data. Sickled cells calculatedinternally.

The results of the modeling and simulations provided in the aboveExamples 8 and 9 for Compound 1 (GBT440) support the use of higher dosesof Compound 1 (e.g., 900 mg, 1200 and 1500 mg) in the treatment of SCD.

7.10 Example 10

The following example describes the making of a Common Blend (CB)capsule formulation at 4.8 kg batch scale.

The CB capsule formulation at 300 mg strength was scaled up to 4.8 kgbatch size and run under GMP conditions to manufacture clinical trialcapsules of Form II of Compound 1 (GBT440). Per the process describedstepwise, 4.114 kg of Form II of Compound 1 and the correspondingquantities of intragranular excipients excluding magnesium stearate werepassed through a 20 mesh screen and added to a high shear granulator andblended for 5 minutes with impellor speed at 300 rpm. The premix wasgranulated by adding water at 60 g/min while mixing at high shear usingimpellor at 300 rpm and chopper at 1200 rpm. After addition of water,the wet granulation was further kneaded or wet massed for 3 min usingimpellor at 300 rpm and chopper at 1200 rpm. The wet granulation wasdried using a fluid bed dryer at an inlet air temperature set at 55° C.and dried until the desired LOD (loss on drying) was attained. The driedgranulation was passed through a co-mill at 1000 rpm to ensure breakingof large agglomerates and to attain a uniform particle sizedistribution.

Extragranular excipient (magnesium stearate) was passed through mesh #40and blended with the granules for 3 minutes at 30 rpm in a V-blender.

Capsules were filled with the final blend using either an semiautomaticor manual encapsulator. The capsules had a an average fill of 350 mggranulation and final capsule weight of approximately 442 mg. 100% ofthe filled acceptable capsules were polished, weight sorted, visuallyinspected for any defects and passed through metal detection prior topackaging.

The capsules were tested by validated analytical methods meeting allproduct quality acceptance criteria, and released for human clinicaluse.

Quantitative compositions of exemplary 300 mg capsules are presented inTable 5, below.

TABLE 5 Quantitative Composition of Exemplary Compound 1, Form IICapsule (300 mg), indicating “Quantity” ((%w/w) and (mg/capsule)),“Function” and “Reference to Standard or Similar” for each component.Quantity Reference Quantity (mg/ to Standard Component (% w/w) capsule)Function or Similar Compound 1 Form II,  85.71% 300.00 Drug In-houseUnmilled substance (intragranular) Hydroxypropyl  4.00% 14.00 Binder USPmethylcellulose (Methocel ® E5 Premium LV) (intragranular)Microcrystalline  3.64% 12.74 Filler NF Cellulose (Avicel ® PH-101)(intragranular) Lactose Monohydrate  2.65% 9.28 Filler NF (ForemostGrade 310) (intragranular) Croscarmellose Sodium  3.50% 12.25Disintegrant Ph. Eur./ (Ac-Di-Sol ®) NF (intragranular) Sterile WaterN/A N/A Granulation USP for Irrigation^(a) Liquid Magnesium Stearate 0.50% 1.75 Lubricant NF (Hyqual ®, Vegetable Source) (extragranular)Total Fill Weight 100.00% 350.02 HPMC (hydroxypropyl N/A 96.0 CapsuleUSP/NF, methylcellulose shell Ph.Eur. (hypromellose)), Swedish orangeopaque, Vcaps ® Plus Coni-Snap, capsules, size 0 Total Weight N/A 446.02

The examples set forth above are provided to give those of ordinaryskill in the art with a complete disclosure and description of how tomake and use the claimed embodiments, and are not intended to limit thescope of what is disclosed herein. Modifications that are obvious topersons of skill in the art are intended to be within the scope of thefollowing claims. All publications, patents, and patent applicationscited in this specification are incorporated herein by reference as ifeach such publication, patent or patent application were specificallyand individually indicated to be incorporated herein by reference.

What is claimed is:
 1. A method for treating sickle cell disease in apatient comprising administering to the patient Compound 1:

wherein the compound is administered in a dose of from about 500 mg/dayto about 1500 mg/day.
 2. The method of claim 1, wherein the compound isadministered in a dose of from about 600 mg/day to about 900 mg/day. 3.The method of claim 1, wherein the compound is administered in a dose ofabout 600 mg/day.
 4. The method of claim 1, wherein the compound isadministered in a dose of about 900 mg/day, or about 1200 mg/day, orabout 1500 mg/day.
 5. The method of claim 1, wherein the compound isadministered in a dose of 600 mg/day.
 6. The method of claim 1, whereinthe compound is administered in a dose of 900 mg/day, 1200 mg/day or1500 mg/day.
 7. The method of claim 1, wherein the compound isadministered once daily.
 8. The method of claim 5, wherein the compoundis administered once daily.
 9. The method of claim 6, wherein thecompound is administered once daily.
 10. The method of claim 1, whereinCompound 1 is a crystalline ansolvate form characterized by at least twoX-ray powder diffraction peaks (Cu Kα radiation) selected from 13.37°,14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ).
 11. The method of claim 10,wherein the crystalline ansolvate form is characterized by at leastthree X-ray powder diffraction peaks (Cu Kα radiation) selected from13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ).
 12. The method ofclaim 10, wherein the crystalline ansolvate form is characterized byX-ray powder diffraction peaks (Cu Kα radiation) at 13.37°, 14.37°,19.95° and 23.92°2θ (each ±0.2°2θ).
 13. The method of claim 10, whereinthe crystalline ansolvate form of Compound 1 is substantially free ofForm I and/or Form N; wherein Form I is characterized by at least threeX-ray powder diffraction peaks (Cu Kα radiation) selected from 12.82°,15.74°, 16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each±0.2°2θ); and wherein Form N is characterized by at least three X-raypowder diffraction peaks (Cu Kα radiation) selected from 11.65°, 11.85°,12.08°, 16.70°, 19.65° and 23.48°2θ (each ±0.2°2θ).
 14. The method ofclaim 5, wherein Compound 1 is a crystalline ansolvate form that ischaracterized by at least two X-ray powder diffraction peaks (Cu Kαradiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ (each±0.2°2θ).
 15. The method of claim 14, wherein the crystalline ansolvateform is characterized by at least three X-ray powder diffraction peaks(Cu Kα radiation) selected from 13.37°, 14.37°, 19.95° and 23.92°2θ(each ±0.2°2θ).
 16. The method of claim 14, wherein the crystallineansolvate form is characterized by X-ray powder diffraction peaks (Cu Kαradiation) at 13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ). 17.The method of claim 14, wherein the crystalline ansolvate form ofCompound 1 is substantially free of Form I and/or Form N; wherein Form Iis characterized by at least three X-ray powder diffraction peaks (Cu Kαradiation) selected from 12.82°, 15.74°, 16.03°, 16.63°, 17.60°, 25.14°,25.82° and 26.44°2θ (each ±0.2°2θ); and wherein Form N is characterizedby at least three X-ray powder diffraction peaks (Cu Kα radiation)selected from 11.65°, 11.85°, 12.08°, 16.70°, 19.65° and 23.48°2θ (each±0.2°2θ).
 18. The method of claim 6, wherein Compound 1 is a crystallineansolvate form that is characterized by at least two X-ray powderdiffraction peaks (Cu Kα radiation) selected from 13.37°, 14.37°, 19.95°and 23.92°2θ (each ±0.2°2θ).
 19. The method of claim 18, wherein thecrystalline ansolvate form is characterized by at least three X-raypowder diffraction peaks (Cu Kα radiation) selected from 13.37°, 14.37°,19.95° and 23.92°2θ (each ±0.2°2θ).
 20. The method of claim 18, whereinthe crystalline ansolvate form is characterized by X-ray powderdiffraction peaks (Cu Kα radiation) at 13.37°, 14.37°, 19.95° and23.92°2θ (each ±0.2°2θ).
 21. The method of claim 18, wherein thecrystalline ansolvate form of Compound 1 is substantially free of Form Iand/or Form N; wherein Form I is characterized by at least three X-raypowder diffraction peaks (Cu Kα radiation) selected from 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andwherein Form N is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 11.65°, 11.85°,12.08°, 16.70°, 19.65° and 23.48°2θ (each ±0.2°2θ).
 22. A capsule dosageform comprising: (i) from about 65% to about 93% w/w of Compound 1; and(ii) from about 2% to about 10% w/w a binder; wherein w/w is relative tothe total weight of the formulation, excluding the weight of thecapsule.
 23. The capsule dosage form of claim 22, further comprisingfrom about 2% to about 10% a disintegrant.
 24. The capsule dosage formof claim 22, further comprising from about 2% to about 10% adisintegrant and about 2% to 35% a filler.
 25. A capsule dosage formcomprising: (i) from about 65% to about 86% w/w of Compound 1; (ii) fromabout 2% to about 6% w/w a binder; (iii) from about 6% to about 25% w/wa filler; (iv) from about 2% to 6% w/w a disintegrant; and (iv) fromabout 0.5% to about 1.5% w/w a lubricant; wherein w/w is relative to thetotal weight of the formulation, excluding the weight of the capsule.26. The capsule dosage form of claim 25 comprising: (i) from about 65%to about 86% w/w of Compound 1; (ii) from about 2% to about 6% w/w abinder; (iii) from about 3.5% to about 25% w/w an insoluble filler, or2.5% to 25% w/w of soluble filler, or 2.5% to 25% of a combination ofsoluble or insoluble filler; (iv) from about 2% to 6% w/w adisintegrant; and (iv) from about 0.5% to about 1.5% w/w a lubricant.27. The capsule dosage form of claim 26 comprising: (i) about 86% w/w ofCompound 1; (ii) about 4% w/w a binder; (iii) about 3.5% w/w aninsoluble filler and 2.5% w/w of soluble filler; (iv) about 3.5% w/w adisintegrant; and (iv) about 0.5% w/w a lubricant.
 28. The capsuledosage form of claim 27 comprising: (i) 85.71% w/w of Compound 1; (ii)4% w/w a binder; (iii) 3.64% w/w an insoluble filler and 2.65% w/w ofsoluble filler; (iv) 2.65% w/w a disintegrant; and (iv) 0.5% w/w alubricant.
 29. The capsule dosage form of claim 27 wherein: Compound 1is a crystalline ansolvate form that is characterized by at least twoX-ray powder diffraction peaks (Cu Kα radiation) selected from 13.37°,14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ); the binder is hypromellose;the insoluble filler is microcrystalline cellulose; the soluble filleris lactose monohydrate; the disintregrant is croscarmellose sodium; andthe lubricant is magnesium stearate.
 30. The capsule dosage form ofclaim 29 wherein the capsule contains 300 mg: 5% of Compound 1, whereincompound 1 is a crystalline ansolvate form that is characterized by atleast two X-ray powder diffraction peaks (Cu Kα radiation) selected from13.37°, 14.37°, 19.95° and 23.92°2θ (each ±0.2°2θ); wherein thecrystalline ansolvate form is substantially free of Form I and/or N;wherein Form I is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 12.82°, 15.74°,16.03°, 16.63°, 17.60°, 25.14°, 25.82° and 26.44°2θ (each ±0.2°2θ); andwherein Form N is characterized by at least three X-ray powderdiffraction peaks (Cu Kα radiation) selected from 11.65°, 11.85°,12.08°, 16.70°, 19.65° and 23.48°2θ (each ±0.2°2θ).